This application relates to a method and kit for the detection of mutans streptococci (S. mutans and S. sobrinus) in saliva specimens.
Many papers have appeared which document the association of mutans streptococci in saliva with dental caries. See, Emilson et al. Scand. J. Dental Res. 93, 96 (1,985); W. Loesche, Microbiol. Reviews 50, 353 (1986; Van Houte et al., Proc. Int. Conf. Cellular, Molecular and Clinical Aspects of Streptococcus mutans, p. 157 (1986, Elsevier).
Mutans streptococci colonize the teeth, and the salivary level of mutans streptococci reflects the number of colonized sites. Emilson et al., Community Dent Oral Epidemiol 10, 111 (1982); Togelius, et al., Acta Odontol. Scand. 42, 157 (1984). Thus, the number of mutans streptococci in saliva can be used for evaluation of caries risk and is useful for monitoring the level of colonization of an individual. Zickert et al. Community Dent. Oral Epidemiol. 10, 77 (1982).
A conventional method for determining the number of mutans streptococci in a saliva sample relies on plating serial dilutions of the sample on a selective medium, incubating the samples to form colonies, counting the number of colonies formed on a manageable plate and back calculating to determine the number of viable bacteria in the original sample. This method, however, is time consuming and requires skilled and accurate handling to achieve reliable results. Further, this method is poorly suited to the development of a test kit.
Another method was described by Kohler et al., J. Clin. Microbiol 9, 584 (1979). In this method, wooden spatulas were exposed to saliva in the mouth of an individual and then placed in contact with an MSB agar surface. MSB agar is selective for growth of mutans streptococci and contains per liter: 10.0 g tryptose, 10.0 g peptone, 1.0 g dextrose, 50.0 g sucrose, 4.0 g K.sub.2 HPO.sub.4, 0.075 g tryptan blue, 0.0008 g crystal violet, 15.0 g agar, 200 units bacitracin and 0.01 g potassium tellurite at a pH of 7.0. The MSB agar plates with the spatula's on the surface were incubated in plastic bags containing expired air from the individual and the number of colonies formed counted.
The Kohler "spatula" method provides a good assessment of the number of mutans streptococci in the mouth and is more convenient than direct plating of saliva samples. Nevertheless, the spatula method is not ideal, in part because of the short shelf life (about seven days) of the MSB agar plates.
It is an object of the present invention therefore to provide a test method for detecting and quantifying mutans streptococci in saliva that makes use of stable reagents that can be readily produced in kit form.